Immediately following transcription heterogenous RNA is complexed with six proteins (A1,A2,B1,B2,C1,C2) that together form a 40S ribonucleoprotein particle that appears to be essential for pre-mRNA packaging, splicing and transport. These six proteins have apparent molecular weights on SDS PAGE that range from 32,000-44,000. All of these "core" hnRNP proteins share the same structural organization consisting of one (type C) or two (type A or B), 90 amino acid residue RNA binding domains at their NH2-termini followed by either a glycine-rich (type A or B) or acidic (type C) domain that contains sites for specific protein:protein interactions. The 90 residue hnRNP binding domain represents a common structural motif found in many other eukaryotic proteins including nucleolin, snRNP associated, sexual differentiation, neural development and poly A binding proteins. In each case this "mobile" RNA binding element is then linked to other domains that are specific to the function of each of the above proteins. The degree of sequence conservation at 21 positions distributed throughout this domain is sufficiently high that it provides on of the few instances where an "activity", RNA binding, can be inferred based solely on an amino acid sequence. Our laboratory and others have isolated cDNA clones and sequenced 5 of the core hnRNP proteins so that we can now address some more interesting and detailed questions. An NMR/X-ray crystallographic approach will be used to solve the three dimensional structure of the 90 residue domain at the NH2- terminus of A1. This prototypical RNA binding domain can be isolated in good yield following limited proteolysis of A1. Functionally important residues within this domain will be identified via a synthetic peptide analogue/in vitro mutagenesis approach. The nucleic acid binding properties of the A1 and type C hnRNP proteins will be examined with particular reference to the role of multiple RNA binding domains in A1 and the possible existence of high affinity chromatography in the absence and presence of synthetic peptide analogues will be used to identify peptide regions that are directly involved in hnRNP protein:protein interactions. In vitro particle reconstitution and nucleic acid binding studies will assess the role of post-translational modifications in hnRNP proteins. These studies are basic to understanding structure/function relationships in a family of eukaryotic, single-stranded RNA binding proteins that are intimately involved in RNA metabolism.